Experimental autoimmune encephalomyelitis (EAE) serves as a useful animal model for organ specific autoimmune disease and is widely used in multiple sclerosis (MS) research. We and others have shown that acquired recognition of new self-antigens, i.e. "epitope spreading," accompanies relapse and chronic progression of EAE and manifests as an orderly sequential appearance of secondary neoautoreactivities as a result of endogenous self-priming. Our goal in the proposed studies is to determine the relative contributions of "primary" (priming), "secondary" (spreading), and "tertiary" (bone marrow regenerating) autoreactive T cells on disease progression. Our recent data indicate that progression of both EAE and MS is accompanied by two concurrent events: 1) the spontaneous regression of the primary established autoreactivity associated with disease onset, and 2) the emergence of the secondary epitope spreading cascade associated with disease progression. The spontaneous regression of primary autoreactivity raises the issue of how long-term memory to self is maintained during autoimmune disease. It is widely believed that long-term memory is due to conventional persistence of the same memory T cells. However, the ability to recall responses to the same self-determinant over time may also be due at least in part to bone marrow regeneration of the same autoreactivity over the course of disease. In aim 1 we will directly address this issue by determining whether bone-marrow regeneration of autoreactivity occurs and whether it recapitulates the pattern of self-recognition observed in the epitope-spreading cascade. Despite the orderly and predictable appearance of epitope spreading in EAE, the relative impact of epitope spreading on disease progression remains in question. In aim 2 we propose to address this issue directly and determine the relative contributions of primary and secondary autoimmune T cell repertoires on progression of EAE by deliberate deletion of primary autoreactivity at various times after EAE onset. Data derived from our proposed studies will indicate whether EAE is perpetuated by repeating cycles of self-recognition cascades and will determine whether long-term memory to self-antigens during autoimmune disease may be due in part to neoregenerated T cells that respond to the same cognate self previously recognized.